The human osteosarcoma cell line U2OS, human normal fibroblast cell line WI38, human normal fibroblast cell line GM00637 and human WRN-deficient fibroblast cell line AG11395 were purchased from the American Type Culture Collection (Manassas, VA, USA). The cell lines were cultured in DMEM medium with 10% fetal bovine serum (FBS; Hyclone Laboratories, Inc., Logan, UT, USA) and grown at 37°C in the presence of 5% CO2.
All predesigned siRNA oligonucleotide duplexes (OnTarget plus option) directed against human XLF or WRN (si-XLF or si-WRN) were purchased from Dharmacon, Inc. (Lafayette, CO, USA). The forward sequences of individual siRNA oligonucleotide duplexes were as follows, for si1-XLF: GCA UUA CAG UGC CAA GTG A dTdT; si2-XLF: CGC UGA UUC GAG AUC GAU UGA dTdT; si1-WRN: CUG UAU CUU CGG GCA CCA A dTdT and i2-WRN: UGA AGA GCA AGU UAC UUG C dTdT. The forward sequence of control siRNA oligonucleotide duplex (si-CTR) was CGU ACG CGG AAU ACU UCG A dTdT.
Mouse monoclonal antibody against β-actin (clone AC15) was purchased from Sigma (St. Louis, MO, USA). Antibodies against XLF (BL3263) and WRN (BL1309) were purchased from the Bethyl Laboratories (Montgomery, TX, USA). Peroxidase-conjugated secondary antibodies were from Jackson ImmunoResearch (West Grove, PA, USA).
All predesigned siRNA oligonucleotide duplexes (OnTarget plus option) directed against human XLF or WRN (si-XLF or si-WRN) were purchased from Dharmacon, Inc. (Lafayette, CO, USA). The forward sequences of individual siRNA oligonucleotide duplexes were as follows, for si1-XLF: GCA UUA CAG UGC CAA GTG A dTdT; si2-XLF: CGC UGA UUC GAG AUC GAU UGA dTdT; si1-WRN: CUG UAU CUU CGG GCA CCA A dTdT and i2-WRN: UGA AGA GCA AGU UAC UUG C dTdT. The forward sequence of control siRNA oligonucleotide duplex (si-CTR) was CGU ACG CGG AAU ACU UCG A dTdT.
Mouse monoclonal antibody against β-actin (clone AC15) was purchased from Sigma (St. Louis, MO, USA). Antibodies against XLF (BL3263) and WRN (BL1309) were purchased from the Bethyl Laboratories (Montgomery, TX, USA). Peroxidase-conjugated secondary antibodies were from Jackson ImmunoResearch (West Grove, PA, USA).